Parkinson’s disease (PD) is a progressive neurological movement disorder characterised by degenerating dopaminergic neurons in the midbrain’s substantia nigra pars compacta (SNpc). In the majority of cases, PD is thought to be caused by a plethora of overlapping factors that combine to cause toxicity in SNpc neurons.
The enzyme nicotinamide N-methyltransferase (NNMT) is expressed at higher levels in the brain of PD patients. However, whether this association is indicative of a causative role in PD is unclear. The biochemical effects of NNMT expression were determined by comparing the NNMT-V5 expressing S.NNMT.LP cell line with its parent SH-SY5Y. Furthermore, the potential for NNMT to potentiate toxicity of β-carbolines (βC), a group of endogenous compounds with increased prevalence in PD, and 4-phenylpyridine (4PP), an environmental analogue of the neurotoxin 1-methyl-4-phenylpyridine, were determined by MTT and ATP assay. Finally, the ability of purified NNMT to metabolise the βC norharman (NH) via 2N-methylation was determined via enzyme assay and the toxicity of 2N-methylated NH (2-MeNH) was determined via MTT and ATP assay.
S.NNMT.LP cells contained a significantly higher ATP content than SH-SY5Y cells (p < 0.01). No difference was seen between S.NNMT.LP and SH-SY5Y toxicity to the βC tetrahydronorharman or 4PP. However, 200 μM NH significantly lowered ATP but not cell viability in S.NNMT.LP cells compared with SH-SY5Y (p < 0.01). NNMT was found to be capable of producing 2-MeNH from NH and S.NNMT.LP cells were significantly protected from 2-MeNH toxicity (p < 0.001). Moreover, 2-MeNH was significantly less toxic to both cell lines than NH (p < 0.05). The metabolism of NH to 2-MeNH may, therefore, be a detoxification mechanism. Accordingly, the expression of NNMT in PD may be a protective response in PD by increasing ATP production and removing NH from the cytoplasm in favour of the less toxic 2-MeNH.
|Date of Award||2015|
|Supervisor||David Barlow (Supervisor) & Richard Parsons (Supervisor)|